EP0133288A2
Expired / Lapsed
EP
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Bacterial detection by nucleic acid hybridization, labeled probes and test kit therefore
Assignee: Bayer Corp
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Filing: 1984-07-25
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Granted: —
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Expires: 2004-07-25
Abstract
Abstract A nucleic acid hybridization method for detecting bacteria in a test sample, such as a mammalian body fluid (e.g., for detecting bacteriuria), employing a polynucleotide probe comprising a base sequence of at least a portion of one of the strands of a tuf or fus gene. Resulting hybridization between the probe and single stranded genomic nucleic acids released from bacteria in the sample indicates bacterial presence and can be detected by employing labeled probes or otherwise detecting duplex formation. The method is particularly applicable to the screening of urine samples for the presence of bacteria belonging to the families Enterobacteriaceae, Psuedomonada- ceae, and Streptococcaceae and particularly those of the genera Escherichia, Klebsiella, Enterobacter, Proteus, Psuedomo- nas, and Streptococcus. The method provides a unique, direct, and rapid means for detecting the presence of bacteria without the need for conventional microbiological culturing.
Inventors
- 👤 Josephine C. Grosch
- 👤 Gary A. Wilson
Bibliographic Details
Application Number
—
Jurisdiction
EP
Data Source
google_patents
Last Synchronized
2026-10-29 12:41
CPC Classifications
C; C12; C12Q; C12Q1/00; C12Q1/68; C12Q1/6876; C12Q1/6888; C12Q1/689; Y; Y02; Y02A; Y02A50/00; Y02A50/30
Claims Analysis
Total Claims: 11
Claim 1
Independent Claim
89 words
A method for detecting bacteria in a test sample, comprising the steps of subjecting the test sample to conditions to release and denature nucleic acids from bacteria present in the sample, contacting the resulting single stranded nucleic acids with a polynucleotide probe under conditions favorable to hybridization between said probe and bacterial base sequences, and determining the presence of hybridized probe, characterized in that the probe comprises a base sequence of at least a portion of one of the strands of a gene selected from tuf and fus genes.
Claim 2
Independent Claim
89 words
A method for detecting bacteria in a test sample, comprising the steps of subjecting the test sample to conditions to release and denature nucleic acids from bacteria present in the sample, contacting the resulting single stranded nucleic acids with a polynucleotide probe under conditions favorable to hybridization between said probe and bacterial base sequences, and determining the presence of hybridized probe, characterized in that the probe comprises a base sequence of at least a portion of one of the strands of a gene selected from tuf and fus genes.
Claim 3
Dependent Claim
14 words
The method of Claim 1 wherein said gene is an E. coli tuf gene.
Claim 4
Dependent Claim
90 words
The method of Claim 1, characterized in that it comprises the steps of: (a) immobilizing the single stranded nucleic acids resulting from treatment of the test sample on a solid support, (b) contacting the solid support with a labeled polynucleotide probe having a base sequence of at least a 10 base portion of one of the strands of a tuf gene sequence, (c) separating unhybridized labeled probe from said solid support, and (d) determining hybridized labeled probe associated with said support by measuring the label comprised in said labeled probe.
Claim 5
Dependent Claim
43 words
The method of Claim 3, characterized in that step (b) involves adsorbing the single stranded DNA resulting from step (a) onto said solid support and thereafter treating the support with foreign nucleic acid to substantially saturate all remaining nucleic acid adsorption sites thereon.
Claim 6
Dependent Claim
46 words
A labeled polynucleotide probe suitable for the method of Claim 1, characterized in that it comprises a label incorporated with a polynucleotide probe comprising a base sequence of at least a portion of one of the strands of a gene selected from tuf and fus genes.
Claim 7
Dependent Claim
22 words
A labeled probe according to Claim 5, characterized in that said homologous base sequence is at least about 10 bases in length.
Claim 8
Dependent Claim
20 words
A labeled probe according to Claim 5 or 6, characterized in that said gene is an E. coli tuf gene.
Claim 9
Dependent Claim
75 words
A test kit suitable for the method of Claim 1, characterized in that it comprises: (a) a solid support capable of immobilizing single stranded nucleic acid resulting from treatment of said test sample to release and denature genomic nucleic acids from bacteria present in said sample, and (2) a labeled polynucleotide probe comprising a base sequence of at least a portion of one of the strands of a gene selected from tuf and fus genes.
Claim 10
Dependent Claim
42 words
A test kit according to Claim 8, characterized in that it additionally comprises foreign nucleic acid for use in substantially saturating nucleic acid adsorption sites on said support after immobilization of said single stranded nucleic acids from bacteria present in said sample.
Claim 11
Dependent Claim
37 words
A test kit according to Claim 8 or 9, characterized in that it additionally comprises a chemical lysing and denaturation agent for treating said test sample to release single stranded genomic nucleic acids from bacteria present therein.
References Cited (Backward Citations)
No citation records indexed.
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